Proteome-Level Analysis of Metabolism- and Stress-Related Proteins during Seed Dormancy and Germination in Gnetum parvifolium.

Gnetum parvifolium is a rich source of materials for traditional medicines, food, and oil, but little is known about the mechanism underlying its seed dormancy and germination. In this study, we analyzed the proteome-level changes in its seeds during germination using isobaric tags for relative and absolute quantitation. In total, 1,040 differentially expressed proteins were identified, and cluster analysis revealed the distinct time points during which signal transduction and oxidation-reduction activity changed. Gene Ontology analysis showed that "carbohydrate metabolic process" and "response to oxidative stress" were the main enriched terms. Proteins associated with starch degradation and antioxidant enzymes were important for dormancy-release, while proteins associated with energy metabolism and protein synthesis were up-regulated during germination. Moreover, protein-interaction networks were mainly associated with heat-shock proteins. Furthermore, in accord with changes in the energy metabolism- and antioxidant-related proteins, indole-3-acetic acid, Peroxidase, and soluble sugar content increased, and the starch content decreased in almost all six stages of dormancy and germination analyzed (S1-S6). The activity of superoxide dismutase, abscisic acid, and malondialdehyde content increased in the dormancy stages (S1-S3) and then decreased in the germination stages (S4-S6). Our results provide new insights into G. parvifolium seed dormancy and germination at the proteome and physiological levels, with implications for improving seed propagation.

A genome for gnetophytes and early evolution of seed plants.

Gnetophytes are an enigmatic gymnosperm lineage comprising three genera, Gnetum, Welwitschia and Ephedra, which are morphologically distinct from all other seed plants. Their distinctiveness has triggered much debate as to their origin, evolution and phylogenetic placement among seed plants. To increase our understanding of the evolution of gnetophytes, and their relation to other seed plants, we report here a high-quality draft genome sequence for Gnetum montanum, the first for any gnetophyte. By using a novel genome assembly strategy to deal with high levels of heterozygosity, we assembled >4 Gb of sequence encoding 27,491 protein-coding genes. Comparative analysis of the G. montanum genome with other gymnosperm genomes unveiled some remarkable and distinctive genomic features, such as a diverse assemblage of retrotransposons with evidence for elevated frequencies of elimination rather than accumulation, considerable differences in intron architecture, including both length distribution and proportions of (retro) transposon elements, and distinctive patterns of proliferation of functional protein domains. Furthermore, a few gene families showed Gnetum-specific copy number expansions (for example, cellulose synthase) or contractions (for example, Late Embryogenesis Abundant protein), which could be connected with Gnetum's distinctive morphological innovations associated with their adaptation to warm, mesic environments. Overall, the G. montanum genome enables a better resolution of ancestral genomic features within seed plants, and the identification of genomic characters that distinguish Gnetum from other gymnosperms.

Melinjo (Gnetum gnemon) Seed Extract Consumption during Lactation Improved Vasodilation and Attenuated the Development of Hypertension in Female Offspring of Fructose-Fed Pregnant Rats.

BACKGROUND: Fructose intake has been correlated with increased prevalence of metabolic disorders including hypertension. In pregnant rats, fructose intake has been reported to have adverse effects on the health of its offspring. This study investigated the effects of gestational maternal fructose consumption and if supplementation with melinjo seed extracts to the maternal diet during lactation could benefit the offspring in later life. METHODS: Pregnant rats were randomly divided into three groups: untreated (CC), fructose-treated (FC), and fructose and melinjo-treated (FM). FC and FM groups received 100 g/L of D(-)-fructose solution by means of the drinking water during gestation while CC received normal drinking water. During lactation, CC and FC groups were given standard commercial laboratory diet, while the FM group was given commercial laboratory diet with 0.1% melinjo seed extracts. After weaning, the offspring were given normal drinking water and standard commercial diet until week 17. The blood pressure of the offspring was monitored until the 16th week. During week 17, the offspring were killed, and the kidneys were collected and analyzed. RESULTS: The level of renal phosphorylated AMP-activated protein kinase (pAMPK) in FM of 17-week female offspring was significantly higher compared with FC and CC groups. Maternal fructose intake down-regulated the renal endothelial isoform of nitric oxide synthetase expression in FC and maternal melinjo seed extract consumption maintained renal endothelial isoform of nitric oxide synthetase expression in FM of 17-week female offspring. In addition, maternal melinjo seed extract intake during lactation lowered the systolic blood pressure in FM of 17-week female offspring. CONCLUSION: Female offspring were more vulnerable to the effects of placental fructose and melinjo seed extracts, suggesting sex-specific sensitivities. In summary, our data show that melinjo seed extract consumption during lactation improved vasodilation and attenuated the development of hypertension in the 17-week female offspring of fructose-fed pregnant rats. Birth Defects Research 110:27-34, 2018. © 2017 Wiley Periodicals, Inc.

Acclimation of seedlings of Gnetum leyboldii Tul. Gnetaceae to light changes in a tropical rain forest / Aclimatación de plántulas de Gnetum leyboldii Tul. Gnetaceae a los cambios de luz en un bosque lluvioso tropical

The neotropical liana Gnetum leyboldii Gnetaceae is a gymnosperm that resembles angiosperms in wood anatomy, overall morphology, and seed dispersal mechanism. Like other woody lianas, seedlings germinate in the shaded forest understory and start climbing towards the canopy, being eposed to sites with etreme differences in light conditions. However, the etent of physiological and structural adjustment to contrasting light conditions in the early regeneration stages of Gnetum is unknown. To answer this question, we analyzed seedling growth and photosynthetic responses using a common garden eperiment with two light regimes: full sun and low light 20 of full sun at La Selva Biological Station, Costa Rica. We also characterized the germination pattern of this species. We monitored one and half-month old seedlings for four months. Leaf structure finely adapted to light treatments, but gas echange properties were buffered by large seed reserves, which dominated biomass distribution about 50 of the total biomass, followed by stem 27, leaf 16 and root biomass 6 across light conditions. The presence of large seeds and the low photosynthetic rates of seedlings in both environments show that G. leyboldii is specialized to eploit deep shade. More research is needed to determine if the patterns found in G. leyboldii are typical of similar lianas that initially eploit deep-shaded understories in their ascension to the canopy.

La liana neotropical Gnetum leyboldii Gnetaceae es una gimnosperma que se asemeja a las angiospermas en la anatomía de la madera, morfología general de la planta y mecanismo de dispersión de semillas. Al igual que otras lianas leñosas, las plántulas se regeneran en el sotobosque bajo dosel cerrado y eventualmente ascienden hacia el dosel, eplotando sitios con diferencias etremas en condiciones lumínicas. Se desconoce el grado de ajuste fisiológico a condiciones lumínicas contrastantes en las primeras fases de regeneración de Gnetum. Para contestar esta pregunta, analizamos las respuestas de crecimiento de las plántulas a ambientes contrastantes de luz de sol y sombra en un jardín común con condiciones de alta cielo abierto y baja luminosidad 20 del ambiente de sol en la Estación Biológica La Selva, Costa Rica. También caracterizamos su patrón de germinación. Monitoreamos plántulas de 1.5 meses de edad por 4 meses. La estructura foliar mostró una fina adaptación a los tratamientos de luz, pero las propiedades de intercambio gaseoso no cambiaron sino que fueron amortiguadas por las reservas de las semillas grandes, las cuales dominaron la distribución de biomasa aproimadamente 50 de la biomasa total seguidas por el tallo 27, la hoja 16 y raíces 6. El tener semillas grandes y plántulas con bajas tasas fotosintéticas muestra que G. leyboldii en su etapa de plántula está adaptado para eplotar la sombra profunda. Se requiere más investigación para determinar si los patrones encontrados en G. leyboldii son típicos de otras lianas que inicialmente eplotan la sombra profunda en su ascensión al dosel.

Acclimation of seedlings of Gnetum leyboldii Tul. (Gnetaceae) to light changes in a tropical rain forest.

The neotropical liana Gnetum leyboldii (Gnetaceae) is a gymnosperm that resembles angiosperms in wood anatomy, overall morphology, and seed dispersal mechanism. Like other woody lianas, seedlings germinate in the shaded forest understory and start climbing towards the canopy, being exposed to sites with extreme differences in light conditions. However, the extent of physiological and structural adjustment to contrasting light conditions in the early regeneration stages of Gnetum is unknown. To answer this question, we analyzed seedling growth and photosynthetic responses using a common garden experiment with two light regimes: full sun and low light (20% of full sun) at La Selva Biological Station, Costa Rica. We also characterized the germination pattern of this species. We monitored one and half-month old seedlings for four months. Leaf structure finely adapted to light treatments, but gas exchange properties were buffered by large seed reserves, which dominated biomass distribution (about 50% of the total biomass), followed by stem (27%), leaf (16%) and root biomass (6%) across light conditions. The presence of large seeds and the low photosynthetic rates of seedlings in both environments show that G. leyboldii is specialized to exploit deep shade. More research is needed to determine if the patterns found in G. leyboldii are typical of similar lianas that initially exploit deep-shaded understories in their accession to the canopy.

Xylem hydraulic and photosynthetic function of Gnetum (Gnetales) species from Papua New Guinea.

Gnetum (Gnetales) species are suggested to be unique extant gymnosperms that have acquired high photosynthetic and transpiration capacities as well as greater xylem hydraulic capacity and efficiency compared with all other extant gymnosperms. This is because Gnetum is the only extant gymnosperm lineage that combines vessels, broad pinnate-veined leaves and an ecological distribution in wet, productive lowland tropical rainforest habitats. Yet, field-based observations on the group's ecophysiological performance are lacking. To test a hypothesis that Gnetum species are ecophysiologically analogous to light-demanding woody tropical angiosperms, stem xylem hydraulic performance, photosynthesis and stomatal conductance were investigated in Gnetum as compared with a diverse group of co-occurring woody plants in a lowland tropical rainforest. It was found that Gnetum species combined low photosynthetic capacity and low stomatal conductances with a low stem water transport ability. The physiological observations are consistent with the general occurrence of Gnetum species in shady, primary forest habitats. These results on Gnetum ecophysiology indicate that the coupling of vessels, broad pinnate-veined leaves and the liana habit do not signal the evolution of a highly opportunistic, light-demanding life history in gymnosperms.

Hydraulic design of leaves: insights from rehydration kinetics.

We examined the leaf hydraulic design in 10 species based on their rehydration kinetics. In all cases, a biphasic response described the temporal pattern of water uptake, with time constants of approximately 30 to 800 s and approximately 800 to 8000 s. The time constants of the fast phase were significantly shorter in the six angiosperms (30 to 110 s) compared with the two single-veined conifer species (>400 s) examined, while the two multi-veined gymnosperm species, Gnetum gnemon and Ginkgo biloba, had time constants for the fast phase of approximately 150 s. Among angiosperm species, the fast phase constituted 50-90% of the total water absorbed, whereas in gymnosperms 70-90% of the water uptake could be assigned to the slow phase. In the four gymnosperms, the relative water uptake corresponding to the fast phase matched to a good degree the relative volume of the venation and bundle sheath extension; whereas in the angiosperm species, the relatively larger water influx during the fast phase was similar in relative volume to the combined venation, bundle sheath extension, epidermis and (in four species) the spongy mesophyll. This suggests a general trend from a design in which the epidermis is weakly connected to the veins (all four gymnosperms), to a design with good hydraulic connection between epidermis and veins that largely bypasses the mesophyll (four of six angiosperms), to a design in which almost the entire leaf appears to function as a single pool.